1. Field of the Invention
This invention relates generally to the field of regulation of gene expression, and more specifically to the modulation of transcription factors and genes in the hedgehog-mediated signaling pathway.
2. Description of the Related Art
Embryologists have long performed experimental manipulations that reveal the striking abilities of certain structures in vertebrate embryos to impose pattern upon surrounding tissues. Speculation on the mechanisms underlying these patterning effects usually centers on the secretion of a signaling molecule that elicits an appropriate response from the tissues being patterned. More recent work aimed at the identification of such signaling molecules implicates secreted proteins encoded by individual members of a small number of gene families. One such family of proteins which may have an influential effect upon patterning activities are those proteins encoded by the hedgehog gene family.
The hedgehog (hh) gene was initially identified based on its requirement for normal segmental patterning in Drosophila (N_sslein-Volhard, C. & Wieschaus, E, Nature 287:795–801, 1980). Its functions include local signaling to coordinate the identities of adjacent cells within early embryonic segments (Hooper, J. E., & Scott, M. P. Early Embryonic Development of Animals, pp. 1–48, 1992) and a later function in cuticle patterning that extends across many cell diameters (Heernskerk, J. & DiNardo, S., Cell, 76:449–460, 1994). The hh gene also functions in the patterning of imaginal precursors of adult structures, including the appendages and the eye (Mohler, J. Genetics, 120:1061–1072, 1988; Ma, et al., Cell, 75:927–938, 1993; Heberlein, et al., Cell, 75:913–926, 1993; Tabata, T. & Kornberg, T. D., Cell, 76:89–102, 1992; Basler, K. & Struhl, G., Nature, 368:208–214, 1994). Genetic and molecular evidence indicates that hedgehog proteins are secreted and function in extracellular signaling (Mohler, J., supra; Lee, et al., Cell, 71:33–50, 1992; Taylor, et al., Mech. Dev., 42:89–96, 1993).
In vertebrates, activities encoded by hh homologues have been implicated in anterior/posterior patterning of the limb (Riddle, et al., Cell, 75:1401–1416, 1993; Chang, et al., Development, 120:3339, 1994), and in dorsal/ventral patterning of the neural tube (Echelard, et al., Cell, 75:1417–1430, 1993; Krauss, et al., Cell, 75:1431–1444, 1993; Roelink, et al., Cell, 76:761–775, 1994).
In most of the embryonic tissues where Hedgehog signaling exerts a patterning effect, activation of the Hedgehog pathway is associated with a proliferative response in target cells. Such embryonic tissues include but are not limited to the developing neural tube, the presomitic mesoderm and the mesoderm of the developing limb bud. In addition, uncontrolled cell proliferation due to inappropriate activation of the Hedgehog signaling pathway is associated with formation of several tumor types including but not limited to basal cell carcinoma, medulloblastoma, and probably breast cancer and glioma. The uncontrolled proliferation in these tumors is probably due to the abnormal activation of transcription factors such as Gli1 that have a normal role in the Hedgehog signaling pathway. For example, in the case of basal cell carcinoma, all or nearly all cases are associated with inappropriately high level expression of the Gli1 transcription factor in basal keratinocytes (Dahmane et al., Nature 1997, 389(6653):876–881). Such inappropriate activation of Gli1 is thought to play a causal role in uncontrolled cell proliferation associated with basal cell carcinoma. The ability to modulate activity of such transcription factors thus represents a possible therapeutic approach to several clinically significant cancers.
The hedgehog polypeptide (HH) is synthesized as a precursor that undergoes autoprocessing to generate an amino-terminal fragment (HH-N) and a carboxy-terminus fragment (HH-C). Lee et al. Science, 266:1528–37, 1994. HH-N contains all the signaling activities of HH, whereas HH-C is responsible for the autoprocessing and attaches a cholesterol molecule to the carboxy-terminal of HH-N to regulate its spatial distribution. (Porter, J. A., et. al. Nature, 374:363–366, 1995. Porter, J. A., et. al. Science, 274:255–259, 1996. Porter, J. A. et. al. Cell, 86:21–34, 1996).